Opioid drugs are the most widely used analgesics in medicine, and are also some of the most widely abused substances. Tolerance to opioids is a major medical problem resulting in reduced efficacy of these compounds for treatment of chronic pain, and increasing the potential for addiction and abuse. In this proposal, we detail an innovative, potentially transformative approach to reveal insight into the mechanisms of opioid tolerance, and propose to develop a humanized, transgenic platform for discovery of small molecules that block or reduce tolerance. Our approach aims to find drugs and drug targets that block tolerance, thereby greatly increasing the efficacy of existing blockbuster pharmaceuticals. Blocking or reducing tolerance to opioids would also be a major step towards preventing dependence. We propose to use the nematode, C. elegans, as a genetic model system to study signaling by a G protein coupled receptor (GPCR) that mediates analgesic and addictive responses to opioids, the -opioid receptor (MOR). We will generate transgenic C. elegans that express human MOR and respond to its stimulation by opioids with behavioral changes. By using a series of cell-specific promoters, we propose to analyze two major types of behavior: chemosensation and locomotion. Further, we will develop assays to study behavioral tolerance caused by prolonged exposure to MOR agonists. In this proposal, we take aim at developing a humanized, transgenic platform to study opioid-induced tolerance with the potential to provide significant and unbiased insight into the mechanisms of tolerance and its chemical attenuation. Because our proposal lets the biology of a living organism provide a solution to the problem of tolerance, our approach is a significant advance over previous efforts that have focused on specific molecular targets using cell-based assays that have, unfortunately, had limited success. The microscopic size of C. elegans and the scalability of the assays we propose to develop are simply not possible in mammalian or vertebrate whole organismal systems further supporting our choice of model system. The successful implementation of our proposal has the potential to be transformative to how drug addiction is studied. Successful implementation of this proposal will lay the ground for the discovery of novel small molecules and genes that regulator of tolerance. The success of our work will encourage the development of humanized C. elegans high throughput behavioral platforms for many other addiction targets including cocaine, methamphetamine, and nicotine.